The lateral superior olive (LSO), a binaural nucleus involved in sound localization, receives converging excitatory and inhibitory projections that are precisely tonotopically matched. The inhibitory projection from the MNTB is refined early in postnatal life through mechanisms that are largely unknown. It was recently found that inhibitory neurons in the MNTB-LSO projection release glutamate during the period of synaptic refinement. The vesicular glutamate transporter VGLUT3, which could support this glutamate release, is transiently expressed in the LSO during the same period and is downregulated shortly after hearing onset. This project will test the hypothesis that VGLUT3 expression in the LSO is downregulated by specific auditory experience. It also aims to test the hypothesis that different vesicular glutamate transporters specifically label the excitatory and inhibitory projections during development. The early glutamatergic transmission in the MNTB-LSO pathway is mediated in large part by postsynaptic NMDA receptors. NMDA receptors have known roles in synaptic plasticity in many systems, with different NMDA receptor subtypes playing distinct roles. Thus, an understanding of the particular NMDA receptor subtypes involved, and whether glutamate is likely to be co-released with GABA and glycine, will be necessary for further studies of glutamate's role in development of the LSO. The third arm of this proposal is to characterize basic physiology of glutamate transmission in both excitatory and inhibitory pathways, as a framework for further studies on the coordinated development of inhibitory and excitatory synapses. Conclusions from these studies will provide insight into the development of a major auditory nucleus, and may ramify beyond the auditory system to shed light on brain disorders involving abnormal inhibition, such as epilepsy and autism.